The rapid detection and differentiation of Mycobacterium tuberculosis complex members from cattle and water buffaloes in the delta area of Egypt, using a combination of real-time and conventional PCR

El-Sayed, Mohamed; Amer, Ali

doi:10.1007/s11033-019-04834-3

Cited by 7 publications

(11 citation statements)

References 30 publications

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“…We found a prevalence of 1.2 and 1.6% tuberculin-positive cases for buffaloes and cattle, respectively, with an overall rate of 1.5%. This is nearly similar to published reports from Egypt 2016 [ 9 ], and lower than the published data at the beginning of 2019 [ 11 ] and the mid of 2019 [ 35 ]. This fluctuation of records may be due to the different times of animal testing and the strict implementation of test and slaughter policy adopted by the General Authority for Veterinary Services for controlling mycobacterium infections in animals and potentially spreading to humans [ 3 ].…”

Section: Discussionsupporting

confidence: 89%

“…Furthermore, the tuberculous lesions in cattle and buffalo carcasses represented 0.7 and 0.45%, respectively from El-Mahalla El-Kubra abattoir, Mid-Delta of Egypt at 2016 [ 10 ]. In April 2019, a prevalence of 3.5% tuberculin-positive cases was declared by Elsayed and Amer [ 11 ], from cattle and Egyptian water buffaloes in Menoufia, Sharkia, Gharbia, Dakahlia, Elbuhaira, and Cairo. During August 2019, a prevalence of 3.43% was confirmed in 27 dairy herds from; Alexandria, Beheira, Daqahlia, Gharbia, Kafr EL Sheikh, Menoufia, and Sharkia [ 12 ].…”

Section: Introductionmentioning

confidence: 99%

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Real-time PCR using atpE, conventional PCR targeting different regions of difference, and flow cytometry for confirmation of Mycobacterium bovis in buffaloes and cattle from the Delta area of Egypt

et al. 2022

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Background Mycobacterium bovis notoriously causes detrimental infections in bovines and humans. In this study, 1500 buffaloes and 2200 cattle were tested by single intradermal comparative cervical tuberculin test and compared with the detection rates of M. bovis isolation, real-time and simplex PCR, and flow Cytometry. Results The tuberculin test is the reference test in Egypt, the positive rate was 54/3700 (1.5%) composed of 18/1500 (1.2%) buffaloes and 36/2200 (1.6%) cattle which were mandatorily slaughtered under the Egyptian legislation, after postmortem examination the non-visible-lesion proportion was 39/54 (72.2%) which surpassed the visible-lesion rate 15/54 (27.8%) with (p < 0.0001). The samples from each case were pooled into one sample representing the case, and the isolation rate of M. bovis was 25/54 (46.3%). Real-time PCR using atpE was positive for mycobacteria on the genus level in 18/18 (100%) and 5/5 (100%) of tissue samples and isolates, respectively; simplex PCR detected M. bovis in 44/54 (81.5%) and 25/25 (100%) of tissue samples and isolates, respectively. Flow Cytometry evaluation of the CD4+, CD8+, WC1+δγ, and CD2+ cell phenotypes showed increased counts in the tuberculin-positive cases compared with negative cases (p < 0.0001), and these phenotypes in the tuberculin-positive cases increased after antigen stimulation than in the negative cases (p < 0.0001). Detection rates of PCR techniques and flow Cytometry exceeded that of bacterial isolation (p < 0.0001) and exhibited a strong correlation. Conclusions The skin test suffers from interference from non-tuberculous mycobacteria able to cause false-positive reactions in cattle and other species. Real-time PCR using atpE, conventional PCR targeting RDs, and flow Cytometry are rapid and accurate methods that correlate with the isolation and can be promising for detection and confirmation of infected live and slaughtered cases.

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Section: Discussionsupporting

confidence: 89%

Section: Introductionmentioning

confidence: 99%

Real-time PCR using atpE, conventional PCR targeting different regions of difference, and flow cytometry for confirmation of Mycobacterium bovis in buffaloes and cattle from the Delta area of Egypt

et al. 2022

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“…The insertion sequence region IS6110 is found in all members of MBTC (Thacker et al., 2011). Being rapid, specific and able to detect small quantities of DNA in clinical samples (Elsayed & Amer, 2019), RT‐PCR is advantageous and was reported to identify 100% of samples (Soliman et al., 2017).…”

Section: Discussionmentioning

confidence: 99%

“…The sequence‐based identification of one Mtb strain from a buffalo bull is interesting and scarce compared to its frequent isolation from cattle (Elsayed & Amer, 2019; Romha et al., 2018). This might point out to a possible role of human in the transmission of such pathogen to bovines via aerosol or sputum‐contaminated ration because cattle‐to‐cattle transmission of such pathogen is minimal (Romha et al., 2018).…”

Section: Discussionmentioning

confidence: 99%

Abattoir survey of bovine tuberculosis in tanta, centre of the Nile delta, with in silico analysis of gene mutations and protein–protein interactions of the involved mycobacteria

Borham

Oreiby

El-Gedawy³

et al. 2021

Transbounding Emerging Dis

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Bovine tuberculosis is a transboundary disease of high economic and public health burden worldwide. In this study, post‐mortem examination of 750 cattle and buffalo in Tanta abattoir, Centre of the Nile Delta, revealed visible TB in 4% of animals and a true prevalence of 6.85% (95% CI: 5.3%–8.9%). Mycobacterial culture, histopathology and RT‐PCR targeting all members of M. tuberculosis complex were performed, upon which 85%, 80% and 100% of each tested lesions were confirmed as TB, respectively. Mpb70‐targeting PCR was conducted on ten RT‐PCR positive samples for sequencing and identified nine Mycobacterium (M.) bovis strains and, interestingly, one M. tuberculosis (Mtb) strain from a buffalo. Bioinformatics tools were used for prediction of mutations, nucleotide polymorphisms, lineages, drug resistance and protein–protein interactions (PPI) of the sequenced strains. The Mtb strain was resistant to rifampicin, isoniazid and streptomycin, and to the best of our knowledge, this is the first report of multidrug resistant (MDR)‐Mtb originating from buffaloes. Seven M. bovis strains were resistant to ethambutol and ethionamide. Such resistances were associated with KatG, rpoB, rpsL, embB and ethA genes mutations. Other mutations and nucleotide polymorphisms were also predicted, some are reported for the first time and require experimental work for validation. PPI revealed more interactions than what would be expected for a random set of proteins of similar size and had dense interactions between nodes that are biologically connected, as a group. Two M. bovis strains belonged to BOV AFRI lineage (Spoligotypes BOV 1; BOV 2) and eight strains belonged to East‐Asian (Beijing) lineage. In conclusion, visible TB was prevalent in the study area, RT‐PCR is the best to confirm the disease, MDR‐Mtb is associated with buffalo TB, and mycobacteria of different lineages carry many resistance genes to chemotherapeutic agents used in treatment of human TB constituting a major public health risk.

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“…The use of techniques and methods involving molecular biology, such as Polymerase Chain Reaction (PCR), has reduced the time required for the diagnosis of bTB. Several studies demonstrate the use of PCR in complementing the identification of M. bovis (Zumárraga et al 2012, Barandiaran et al 2019, Fico et al 2019, such as the work of Elsayed & Amer (2019), who used PCR directly from tissue samples, demonstrating the technique's speed compared to cultivation for species identification. Sales et al (2014) compared different primers for the detection of M. bovis, proving that Mb.400 primers, targeting fragments flanking the region of differentiation 4 (RD4), are the ones that demonstrated the best results in the PCR for bTB.…”

Section: Discussionmentioning

confidence: 99%

Bovine tuberculosis in safari park in Brazil

et al. 2021

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Bovine tuberculosis (bTB) is an infectious disease caused by Mycobacterium bovis, affecting domestic animals, wild animals and humans. In captivity, for wild animals, bTB represents a risk to animal keepers and zoo visitors, in addition to the possibility of spreading the infection to domestic animals or through the trade of infected wild animals. Sambar (Cervus unicolor), red deer (Cervus elaphus) and fallow deer (Dama dama) from a safari park in the State of Rio Grande do Sul, Brazil, showed a clinical condition of dyspnea and weight loss. Some animals died and showed lesions suggestive of tuberculosis (LST), which were confirmed by histopathology. After the interdiction of the safari park by the state veterinary authorities, 281 deer were euthanized with the authorization of the “Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renováveis” (IBAMA). Retropharyngeal and submandibular lymph nodes and viscera were collected from 21 animals, which were grown in Stonebrink medium for up to 90 days. After DNA extraction from the bacterial colonies, PCR was performed for targets flanking the region of differentiation 4 (RD4). Of the 21 samples, 14 (66.7%) presented LST with a granulomatous appearance, a whitish coloration, and caseous or calcified consistency, and seven samples (33.3%), showed no lesions. In the culture of 14 samples with LST, 13 (92.8%) presented bacterial growth compatible with M. bovis. In the cultivation of the seven samples without LST, four (57.1%) presented colonies compatible with M. bovis. PCR and DNA sequencing of the PCR amplicons detected as positive all the 17 (100%) bacteriological cultures suggestive of M. bovis, thus confirming the outbreak of bTB in deer. Decisions about positive tested and suspicious animals should be taken based on the evaluation of the risk of transmission to the rest of the zoological animals, animal welfare, conservation considerations and, the zoonotic potential of this pathogen.

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The rapid detection and differentiation of Mycobacterium tuberculosis complex members from cattle and water buffaloes in the delta area of Egypt, using a combination of real-time and conventional PCR

Cited by 7 publications

References 30 publications

Real-time PCR using atpE, conventional PCR targeting different regions of difference, and flow cytometry for confirmation of Mycobacterium bovis in buffaloes and cattle from the Delta area of Egypt

Real-time PCR using atpE, conventional PCR targeting different regions of difference, and flow cytometry for confirmation of Mycobacterium bovis in buffaloes and cattle from the Delta area of Egypt

Abattoir survey of bovine tuberculosis in tanta, centre of the Nile delta, with in silico analysis of gene mutations and protein–protein interactions of the involved mycobacteria

Bovine tuberculosis in safari park in Brazil

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